Mechanisms of membrane estrogen receptor-α-mediated rapid stimulation of Ca2+levels and prolactin release in a pituitary cell line

Abstract

The role of membrane estrogen receptor-α (mERα) in rapid nongenomic responses to 17β-estradiol (E2) was tested in sublines of GH3/B6 rat prolactinoma cells selected for high (GH3/B6/F10) and low (GH3/B6/D9) mERα expression. E2elicited rapid, concentration-dependent intracellular Ca2+concentration ([Ca2+]i) increases in the F10 subline. Lack of inhibition by thapsigargin depletion of intracellular Ca2+pools, together with abrogation of the response in Ca2+-free medium, suggested an extracellular source of Ca2+for this response. The participation of voltage-dependant channels in the E2-induced [Ca2+]iincrease was confirmed by the specific L-type Ca2+channel inhibitor nifedipine. For comparison, the D9 mERα-depleted subline was insensitive to steroid action via this signaling mechanism. [Ca2+]ielevation was correlated with prolactin (PRL) release in the F10 cell line in as little as 3 min. E2caused a much higher PRL release than KCl treatment (which caused maximal Ca2+elevation), suggesting that secretion was also controlled by additional mechanisms. Participation of mERα in these effects was confirmed by the ability of E2-peroxidase (a cell-impermeable analog of E2) to cause these responses, blockage of the responses with the ER antagonist ICI 182 780, and the inability of the E2stereoisomer 17α-E2to elicit a response. Thus rapid exocytosis of PRL is regulated in these cells by mERα signaling to specific Ca2+channels utilizing extracellular Ca2+sources and additional signaling mechanisms.